The major focus of our research is to understand how chromatin regulatory mechanisms influence nuclear and epigenetic programs, and how de-regulation of these mechanisms contributes to aging and disease. SIRT6 is a chromatin regulatory factor in the sirtuin family of enzymes. SIRT6-deficiency in mice leads to shortened lifespan and phenotypes associated with aging, cancer, and metabolism. Conversely, SIRT6 over- expression in mice can protect against metabolic disease and extend lifespan. Thus, studying SIRT6 function promises to elucidate fundamental molecular mechanisms that underlie healthy aging and longevity. Previously, we showed that SIRT6 selectively regulates specific chromatin marks associated with epigenetic and gene-regulatory functions. We linked chromatin regulation by SIRT6 to key nuclear processes that impact on aging and cancer, including telomere maintenance, DNA repair, and aging-associated gene expression changes. Here, we focus on new functions of SIRT6 in chromatin silencing mechanisms that are deregulated in aging and age-related biology. We propose molecular, genomic, and functional studies to elucidate the role of SIRT6 in maintaining heterochromatin silencing at non-coding regions of the genome, and ask how deregulation of this silencing can lead to cellular dysfunction associated with aging. In Aim 1, we will study the molecular mechanisms of SIRT6 in heterochromatin silencing of repetitive DNA elements at mammalian centromeres. Defects in heterochromatin maintenance at centromeres are observed in the contexts of both aging and cancer. We will characterize the biochemical activity of SIRT6 at centromeric repeats, how this is regulated during mitosis, and how it affects higher order chromatin changes. These studies should provide insights into the dynamics of histone acetylation that drive higher order chromatin changes relevant for aging and cancer biology. In Aim 2, we will characterize the role of SIRT6 in heterochromatin maintenance in specific aging and cancer-related physiologic settings. These include human cancer cell types, where SIRT6 has tumor suppressive functions, and primary mammalian cells, where SIRT6 guards against cellular senescence and regulates age-dependent changes in epigenetic plasticity. We will also investigate heterochromatin defects at another class of repetitive sequences that are de-regulated in aging, endogenous transposable elements. In Aim 3, we will characterize the functional effects of heterochromatin maintenance by SIRT6 on cellular homeostasis. We will examine how chromatin defects at centromeres leads to mitotic chromosome segregation defects, and investigate the hypothesis that these defects promote genomic instability and cellular senescence. We will also examine the mechanisms through which heterochromatin loss at transposable elements can promote genomic instability. Together, these studies should provide insights into fundamental chromatin mechanisms in aging biology.